Microbiology 6- Prevention and treatment of viral diseases

Question 1

What are prophylactics and antivirals used for

Answer 1

We use drugs and vaccines to combat viruses. Most vaccines are used prophylactically. Most antiviral drugs are given after the person is infected as a therapeutic agent, although in controlling diseases outbreaks prophylactic antiviral administration has been used.

Question 2

Describe the difference between prophylaxis and antivirals

Answer 2

Prophylaxis is preventing disease before the aetiologic agent is acquired, by vaccination or giving drug before infection.
Therapy is treating the disease after the host has been infected.

Question 3

Describe vaccines

Answer 3

Type of prophylaxis, used as a public health measure, not specific to the patient, often designed for target groups, prevent the spread of infection through herd immunity,

Question 4

How can vaccines have a negative effect

Answer 4

They can prime the immune system to respond badly when virus enters

Question 5

Describe the characteristics of viruses

Answer 5

Prophylactic

Live or inactive

Herd immunity or defined target group?

Safety > efficacy (RSV vaccine in 1970s!)

Governments and WHO

Question 6

Describe the characteristics of antiviral therapy

Answer 6

Therapeutic

Random screen or rational design

Define target group: very sick or over the counter?

Individuals

Question 7

Describe how smallpox was eradicated

Answer 7

No animal reservoir
No latent or persistent infection
Smallpox was an easily recognised disease
The vaccine was effective against all strains of virus
Vaccine properties. Potency, low cost, abundance, heat stability, easy administration
WHO determination
$250 million

Question 8

List some examples of live attenuated vaccines

Answer 8

Rotavirus

Rubella

Smallpox

Varicella

Yellow fever
Polio
MMR
Adenovirus
Influenza

Question 9

List some viruses targeted by inactivated vaccines

Answer 9

Hep A
Polio
Rabies
Tick-borne encephalitis
Japanese encephalitis

Question 10

List some viruses targeted by purified subunit vaccines

Answer 10

Influenza

Question 11

List some viruses targeted by cloned subunit vaccines

Answer 11

Hep B

Human papilloma virus

Question 12

Why is there a possibility to have vaccines other than live attenuated

Answer 12

Replication of the virus is not required to trigger the immune response, hence the whole virus is not needed. We just need their shapes, this allows for fractionated viruses to be utilised, which is a subunit of the virus. As well as inactivated vaccines, where the virus has been inactivated by heat or chemical treatment to prevent it from replicating.

Question 13

Describe the different applications of cloning a virus in vaccination

Answer 13

With knowledge of the viral genome, we can use genetic engineering to incorporate it into attenuated viral vectors- will trigger an immune response.
We can inject the viral DNA into muscle cells, immune response against own body cells.
Or we can use lettuce leaves or yeast as a vector- eat it- immune response

Question 14

Describe how a virus can be attenuated to generate a live attenuated vaccine

Answer 14

The pathogenic virus is isolated from a human patient and grown in human cultured cells
The cultured virus is used to infect monkey cells
The virus acquires many mutations to allow it to grow well in monkey cells
The virus no longer grows well in human cells and maty be a candidate for a vaccine.

Question 15

Describe the pros and cons of live attenuated vaccines

Answer 15

Rapid broad, long lived immunity
Dose sparing
Cellular immunity

BUT
Requires attenuation
May revert

Question 16

Describe the pros and cons of inactivated vaccines

Answer 16

Safe
Can be made from wild type virus

BUT
Frequent boosting required
High doses needed

Question 17

Describe Examples of viruses for which both live and inactivated vaccines are available

Answer 17

Influenza
Inactivated virus or HA subunit
Updated regularly
DOES NOT give the recipient the flu!!!

LAIV is cold adapted
FluMist delivered intranasally
Updated regularly
Introduced for children in the UK 2013

Poliovirus
Salk inactivated vaccine

Sabin live attenuated vaccine
1 in 7 million vaccinations associated with poliomyelitis
Persisting in immunosuppressed individuals
Salk will be required for the ‘end game’.

Question 18

Describe the rotavirus vaccine

Answer 18

Rotarix is a live attenuated rotavirus reassortant virus.
In the developing world it can massively reduce deaths due to rotavirus infection that leads to dehydration from vomitting and diarrhoea.
In the UK 1 in every 50 babies will be admitted to hospital because of rotavirus infection in the first 5 years of life.

Early use of the vaccine in the US highlighted that it can cause intussusception (bowel blockage) in older babies ( older than 3 months). Thus this vaccine is only given to babies less than 15 weeks in the UK.

Question 19

Describe the shingles vaccine

Answer 19

Shingles is a painful rash resulting from the reactivation of a latent varicella zoster virus infection (chicken pox).
Shingles occurs in people after stress, and is more common and more serious in the elderly as their immune system wanes.
Even after the rash has gone (7-10days) pain can remain as Post Herpetic Neuralgia (PHN).
The live attenuated vaccine is similar but distinct from the chick pox vaccine given to children in some countries (not routinely given in UK).
Shingles vaccine introduced in September 2013 and available today but only for those aged 70 or 78.

Question 20

Describe some subunit vaccines

Answer 20

Hepatitis B virus
sAg of HBV cloned and expressed in yeast
Papillomavirus
Virus Like Particles from recombinant coat proteins
Gardasil and Cervarix

Question 21

What type of particles does the immune system respond better to

Answer 21

Multi-valent particles (viruses that express a lot of their antigen receptors). Although viruses are not injected, they look like viruses but cannot replicate or cause disease

Question 22

When do we see the better effects of attenuation

Answer 22

When the virus is exposed to new tissue- forward genetic procedure. Don’t know which mutations will develop, or what the key mutation will be.

Question 23

Why do live vaccines only need a small dose

Answer 23

They replicate inside the cell

Question 24

What can over attenuation cause

Answer 24

May develop the disease the vaccine was intended to protect.

Question 25

Describe the ebola vaccines

Answer 25

GSK: Chimpanzee adenovirus vectored vaccine that expresses Ebola G protein

Merck : Vesicular Stomatitis Virus vectored vaccine expressing Ebola virus G protein.

Question 26

Describe the passive immunization methods of treating ebola

Answer 26

Zmapp :
A cocktail of 3 monoclonal antibodies raised against G protein of a previous strain of Ebola virus given as passive immunotherapy
Unusually production in plants: limited capacity.

Serum therapy: blood from survivors

Question 27

Describe antiviral treatment

Answer 27

Interferons to induce the hosts natural antiviral response
Drugs with specific antiviral activity

Treatment that alleviate symptoms but do not inhibit virus replication

Question 28

Describe the interferon system

Answer 28

Cells recognise foreign material inside cell. Interferon production increased, acts on neighbouring cells to warn them that a virus is present, increased transcription of interferon stimulating genes- protection against virus.

Question 29

What is meant by pegylation

Answer 29

A chemical modification which allows the protein to survive longer inside the body in terms of its pharmacokinetics- impaired pharmacokinetic performance

Question 30

Why did recombinant interferons not work

Answer 30

Recombinant interferon 1981-1982
Side effects due to induction of cytokines and other ISGs- stimulated inflammatory response (fever, aches) which made patients feel worse
Was a few years ago the only option for some viruses, eg. Pegylated IFN given with ribavirin for hepatitis C virus.

Question 31

Why are viruses hard to kill

Answer 31

NO peptidoglycan cell wall
No ribosomes
No cell membranes
Viruses are intracellular obligate parasites.
This makes them particularly difficult to combat with chemotherapeutic agents.
It is hard to find a stage of the virus replication cycle to attack with a drug that does not involve a host function. A drug that inhibited viral translation for example would knock out our own cells’ ability to translate mRNAs.

Question 32

Describe the targets for antiviral drugs

Answer 32

In reality most antiviral drugs target viral enzymes, often acting as substrate analogues, or are cancer like drugs- nucleoside analogues.

Increased understanding of structure of viral components and enzymes can lead to rational drug design.
Most antiviral drugs used today are very specific for the particular virus their work against. They usually target viral enzymes that have been found to differ from any enzymes used by our own
cells. They are difficult to use because an accurate diagnosis is required to inform the correct drug choice. Viruses also often develop resistance to the drugs particularly if they are used individually.

Question 33

Describe acyclovir action and specificity

Answer 33

modified nucleoside incorporated into DNA
lack of 3’ -OH prevents phosphodiester bond formation
Only activated inside virus infected cells
Higher affinity for viral DNA polymerase than for host cell polymerase
Resistance is rare but maps to thymidine kinase.

Question 34

Describe how we can target HIV-1

Answer 34

Targeting HIV-1. HIV-1 life cycle and classes of antiretroviral agents that interfere with these specific steps. The seven steps in the HIV lifecycle are identified by numbered circles. Classes of antiretroviral drugs are shown as red lines near the life cycle step that they inhibit. NNRTI, non-nucleoside reverse transcription inhibitor; NRTI, nucleoside reverse transcription inhibitor; RT, reverse transcription; RTI, reverse transcription inhibitor.

Question 35

Describe the roles of fusion and entry inhibitors

Answer 35

Binds CCR5

Binds gp41

Question 36

Describe the roles of non-nucleoside reverse transcriptase inhibitors

Answer 36

Inhibit RT conversion of
viral RNA to DNA
Block allosteric site on RT
Binds gp41
SE: rash

Question 37

Describe the roles of nucleoside/nucleotide analogues

Answer 37

Inhibit RT
Drug incorporated into 
growing vDNA strand
causing early termination
SE: lactic acidosis
Hepatic steatosis

Question 38

Describe the roles of integrase inhibitors

Answer 38

Inhibit insertion of
pro-viral DNA into 
host genome.
SE: insomnia 
depression

Question 39

Describe the roles of protease inhibitors

Answer 39

Inhibit formation of mature viral particles.
SE: hyperlipidemia
GI intolerance

Question 40

Explain how we currently treat HIV

Answer 40

Zidovudine AZT is also a nucleoside analogue. It was the first anti HIV drug but resistance quickly emerged. Current HIV therapy uses three or four different drugs in combination known as HAART highly active antiretroviral therapy. This prevents the virus from being able to generate resistance mutants but does lead to difficult drug regimens and is associated with significant side effects.

Question 41

Describe antiviral therapy for Hep C

Answer 41

HCV is a hepatotropic flavivirus that was spread widely in the 1970s in blood products before screening was put in place.
170 million people are chronically infected and 4% will proceed to hepatocellular carcinoma.
For more than 20 years therapy relied on interferon treatment with ribavirin.
This was only effective (produced a sustained virological response SVR) in 50% patients infected with the most common genotype 1 of the virus, and carried unpleasant side effects.

Question 42

Describe direct acting antivirals against Hep C

Answer 42

Glecaprevir, grazoprevir,simeprevir, paritaprevir, and voxilaprevir are HCV protease inhibitors that offer several benefits over earlier protease inhibitors (such astelaprevirandboceprevir), including fewer drug-drug interactions, improved dosing schedules, and less frequent and less severe side effects.
Sofosbuviris a nucleoside polymerase inhibitor with high potency across all six genotypes, a very high barrier to resistance. Unlike other candidate drugs in this class, it is generally well tolerated.
Dasabuvir is a non-nucleoside polymerase inhibitor, less potent and lower threshold for resistance than other classes; main role as an adjunct to more potent compounds with higher barriers to resistance.
Agents that inhibit NS5A are generally quite potent and are effective across genotypes, but have a low barrier to resistance and variable toxicity profiles.
Several direct-acting antivirals are only available as part of fixed-dose combinations. These areelbasvir-grazoprevir,glecaprevir-pibrentasvir,ledipasvir-sofosbuvir,sofosbuvir-velpatasvir,sofosbuvir-velpatasvir-voxilaprevir, andombitasvir-paritaprevir-ritonavir, the latter of which is administered with dasabuvir for certain genotypes.
These drugs were made possible due to knowledge of structures and functions of viral proteins and enzymes

Question 43

How do we treat Hep C

Answer 43

Interferon is also currently used in combination with ribavirin to treat HCV.

Question 44

Describe antivirals targeting influenza replication

Answer 44

Most drugs previously used can no longer be used to due to the development of resistance.
Neuraminidase inhibitors are antivirals still used (zanamivir and oseltamivir).

These agents target neuraminidase, which is responsible for cleaving the bonds between the emerging virus and the cell, therefore freeing the virus to penetrate respiratory secretions and replicate.

Question 45

Explain how NA inhibitors work

Answer 45

Competitive inhibitors, irreversible bind to binding site of NA. Stay there due to interactions with chemical pocket (negatively charged), inhibitors have highly positively charged groups. Need to know crystal structures of the substrate.

Question 46

List and compare the strategies underlying the search for novel antiviral agents

Answer 46

A new era of antiviral drug discovery has been sparked by the genomics revolution. We can now identify many host cell genes that viruses need for their replication The hope is that we can target some of them because our genome has some redundancy.
A good example of this is the CCR5 protein used by HIV to enter cells and the fact that a group of exposed uninfected individuals remain resistant to HIV because they have a 32bp deletion in CCR5 but are otherwise completely healthy.
There is hope that more broadly acting antivirals may be discovered. New breakthroughs include small molecules that freeze the lipids on enveloped viruses but seem not to affect the plasma membranes of our own host cells.

Question 47

What is a golden target for antivirals

Answer 47

Protease.

Question 48

Describe cold-adapted LAIV

Answer 48

Can only replicate in 34 degree environment of the nose, mutations which prevent replication in the body